N-acetylneuraminic acid (N-acetyl-D-neuraminic acid, Neu5Ac) is an important milk powder additive, which can improve immunity of infants [1], meanwhile it can be used as a precursor for synthesizing anti-influenza A/B type virus drugs [2]. Synthesizing Neu5Ac by using ManNAc and pyruvic acid as the substrates under catalysis by Nal is the currently most primary synthesis route for Neu5Ac. Nal has been used in industrial synthesis of Neu5Ac [3-5], and synthesis of Neu5Ac from ManNAc and pyruvic acid under catalysis by Nal is a reversible reaction. Nal is widely distributed in the nature, and it is found in bacteria and mammals [6]. Many pathogenic bacteria, after invasion into human body, decompose human Neu5Ac by Nal as their carbon source and nitrogen source [6], thus currently there are a great number of reports on Nal from pathogenic bacteria. Besides Nal derived from pathogenic bacteria, there is also Nal found from food safe (generally regarded as safe, GRAS) strains, such as Lactobacillus plantarum WCFS1 [7] and Taphylococcus carnosus TM300 [8]. Because the substrate pyruvic acid is cheap [9] and Nals have relatively high temperature stability [10], it has been widely applied in the synthesis of Neu5Ac. However, currently all of Nal have a common defect: in a reversible catalytic synthesis reaction of Neu5Ac, the Nal is more prone to decompose Neu5Ac [7, 8, 11-14].
To obtain Nal of high activity and to resolve the issue of the chemical equilibrium of Nal being prone to decomposing Neu5Ac, this patent cloned and expressed N-acetylneuraminic acid aldolase (CgNal) from food safe strain Corynebacterium glutamicum ATCC 13032 [15]. It belongs to one of Nal family, comprising 312 amino acids, and its accession number in Genbank is NP_601846, its amino acid sequence is shown in SEQ ID NO: 2. The gene encoding this protein comprises 939 bp bases, its accession number in the Genbank is NC_003450.3, and its gene sequence is shown in SEQ ID NO:1. Reports on using CgNal in Neu5Ac synthesis has not been found until now.